Introduction

Silage production is essential to many agricultural production systems. It is the process of converting forage to a nutritional, storable feed to use in either feed mixes (in a dairy or feedlot), or as a safeguard for times of drought. Silage is produced primarily by the process of fermentation and has a variety of benefits when compared with forage production for hay. These include; retained nutritional value, and a smaller time needed to be designated for wilting (Moran, J 2005).

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Although the process of silage production seems relatively simple, there are still a variety of ways in which it can go wrong (Bernardes et al. 2018). The factors that have the most influence on overall silage quality are; stage of maturity, carbohydrate content, contact to air and moisture content (Bernardes et al. 2018). This is due to the biochemical pathways which occur during the fermentation process. The ensiling process has various biochemical stages.  The first of these being the aerobic stage where air is still present in the silage (Moran, J 2005). At this stage the oxygen trapped in the sealed silage is eliminated through the respiration of the plant matter as well as the aerobic activities of yeasts and bacteria (Moran, J 2005). The fermentation phase begins once there is no more oxygen in the sample. In this phase the lactobacillus bacteria convert pyruvate and available carbohydrates to lactic acid and an increase in acidity cause the pH of the silage to drop. Sugars were added to samples in this experiment so as to show the effect of increased carbohydrate availability on lactic acid production to produce a quality silage. Once this process has been completed, the silage pH becomes stable between 3.7 and 4.2 assuming no more air or water is able to enter. If water or air were to come into contact with the sample, the aerobic spoilage phase would occur (Moran, J 2005). Aerobic spoilage occurs due to the breakdown of preserving organic acids by yeasts and acetic acid bacteria. This causes the pH to rise and can lead to a second stage of spoilage consistent with microorganisms, moulds and enterobacteria (Moran, J 2005).

When silage is successfully produced it has a pH range of 3.7 – 4.2 (Ward & Ondarza 2008). When the pH is higher or lower than this range, it becomes too acidic for livestock, or not acidic enough to prevent microbial attacks (Ward & Ondarza 2008, Gibson, T 1965). The microorganism Clostridia is one example of this. Clostridia is found in fresh forage and is able to break down protein and produce butyric acid in the presence of oxygen or at high pH levels (Zheng et al. 2017). This makes the silage unstable and susceptible to changes (Zheng et al. 2017). Silage affected by Clostridia usually have higher silage dry matter losses, reduced energy levels and palatability, and high ammonia nitrogen content (Ward & Ondarza 2008, Gibson, T 1965). These characteristics are not desirable for livestock feed. 

Grass type also plays a role in the outcome of a silage sample (Kaiser et al. 2004). Usually silage is produced from temperate (C3, ryegrass, white clover – a C3 legume) forages. These forages have high leaf material and free water-soluble carbohydrates (Kaiser et al. 2004). However, in warmer areas tropical (C4, kikuyu) grasses are often prominent and therefore must also be considered for silage production (Kaiser et al. 2004). Tropical grasses are often found to have a higher moisture content than temperate grasses (Hattersley, P 1983). This means the wilting process is exceptionally important for silage made from these grasses to prevent butyric acid formation as well as effluent production (Rodriguez et al. 1989). As well as this, tropical grasses tend to have a lower concentration of water-soluble carbohydrate than that of the temperate grasses. This means ensiling can be significantly impaired (Bernardes et al. 2018).

Materials and Methods

Kikuyu, ryegrass and clover were harvested and cut into 1 – 1.5 cm sections and these samples were sealed into separate bags. Thick stems and non-grass components were removed and four of these bags were vacuum sealed, while two had no vacuum applied and were sealed with air still inside. Each bag weighed 300-400g, table 1 shows the percentage sugar applied to the samples in each bag.

Table 1: bag contents and seal type

% sugar added

Type of seal

Kikuyu

Fresh

N/A

0% sugar

Vacuumed

1% sugar

Vacuumed

2% sugar

Vacuumed

3% sugar

Vacuumed

0% sugar

Non-vacuumed – heat sealed

3% sugar

Non-vacuumed – heat sealed

Ryegrass

Fresh

N/A

0% sugar

Vacuumed

1% sugar

Vacuumed

2% sugar

Vacuumed

3% sugar

Vacuumed

Clover

Fresh

N/A

0% sugar

Vacuumed

1% sugar

Vacuumed

2% sugar

Vacuumed

3% sugar

Vacuumed

The silage samples were stored for 28 days at room temperature.

An analysis of the silage treatments was completed using a pH probe as well as titrations to measure; nitrogen, ammonia, VFAs and lactic acid.

These measurements were analysed to find the levels of dry matter, organic matter, ammonia and fermentation acids present

Results

Table 2: Percentage of lactic acid and VFAs (lactic, acetic, propionic and butyric) on a Dry Matter basis within each treatment. Buffering capacities (mmol NaOH used to titrate from pH 3 to 7) are also included.

Forage type

Fresh or silage

vacuum or non vacuum

pH

% Lactic Acid (DM)

% Acetic Acid (DM)

% Propionic Acid (DM)

% Butyric Acid (DM)

Buffering capacity         mmol NaOH used to titrate from pH 3 to 7

Kikuyu

Fresh

n/a

6.4

0.012

0

0

0.028

0.95

 0%

Silage

non vacuum

6.8

0.095

0.261

0.043

1.091

1.1

 3%

Silage

non vacuum

5.9

1.012

0.728

0.029

1.540

1.9

 0%

Silage

vacuum

6

1.269

1.365

0.032

1.215

2.25

 1%

Silage

vacuum

5

3.396

0.893

0.029

1.383

2.15

 2%

Silage

vacuum

4.8

4.404

0.764

0.006

0.242

2.3

 3%

Silage

vacuum

4.3

5.995

0.669

0.012

0.068

2.55

Ryegrass

Fresh

n/a

6.4

0.014

0

0

0

0.7

 0%

Silage

vacuum

4.6

7.195

0.953

0.026

0.624

2.35

 1%

Silage

vacuum

4.4

7.918

0.968

0.044

0.451

2.45

 2%

Silage

vacuum

4.3

8.344

0.466

0.042

0.324

2.65

 3%

Silage

vacuum

4.3

6.879

0.541

0.035

0.316

2.4

Clover

Fresh

n/a

6.1

0.016

0

0

0

1.05

 0%

Silage

vacuum

4.2

7.251

0.556

0.092

0.327

2.4

 1%

Silage

vacuum

3.9

7.580

0.473

0.076

0.342

2.5

 2%

Silage

vacuum

3.9

8.689

0.408

0.068

0.275

2.3

 3%

Silage

vacuum

4

6.855

0.440

0.071

0.517

2.3

Graph 1: buffering capacities of fresh grasses showing amount of NaOH needed to titrate from pH 3 to 7.

Graph 2: buffering capacities of 0% sugar silages showing amount of NaOH needed to titrate from pH 3 to 7.

Graph 3: buffering capacities of 1% sugar silages showing amount of NaOH needed to titrate from pH 3 to 7.

Graph 4: buffering capacities of 2% sugar silages showing amount of NaOH needed to titrate from pH 3 to 7.

Graph 5: buffering capacities of 3% sugar silages showing amount of NaOH needed to titrate from pH 3 to 7.

Table 3: Nutritional values of each silage treatment

Forage type

Fresh or silage

vacuum or non vacuum

Carbohydrate added (%)

 % Dry Matter (DM)

% Organic matter (OM)

% Ash

% Crude Protein       (DM basis)              (N x 6.25)

Ammonia N as a % of total N

Kikuyu

Fresh

n/a

n/a

23.2

91.9

8.1

18.2

0

Silage

non vacuum

0

24.1

90.1

9.9

18

8.7

Silage

non vacuum

3

23.9

89.2

10.8

17.8

12.4

Silage

vacuum

0

23.8

91.1

8.9

18.3

8.9

Silage

vacuum

1

24

90.8

9.2

18.2

6.3

Silage

vacuum

2

24.1

91

9

18.2

3.6

Silage

vacuum

3

24.3

90.8

9.2

17.9

3.1

Ryegrass

Fresh

n/a

n/a

18.1

90.5

9.5

21.3

0

Silage

vacuum

0

18.2

89.1

10.9

21.5

7.2

Silage

vacuum

1

18.4

89.5

10.5

20.9

6.1

Silage

vacuum

2

18.4

89

11

21.1

4.1

Silage

vacuum

3

18.4

88.9

11.1

21.5

6.1

Clover

Fresh

n/a

n/a

17.5

88.2

11.8

22.6

0

Silage

vacuum

0

17.9

86.9

13.1

22.9

7.9

Silage

vacuum

1

18

86.2

13.8

23.1

6.8

Silage

vacuum

2

18

87.1

12.9

22

6.2

Silage

vacuum

3

17.9

87.2

12.8

22.9

5.1

Discussion

After a careful analysis of the biochemical and nutritional compounds found in each of the samples, it was found that both air and added sugar content played a significant role in the successfulness of a particular silage. The affects these components had on the production of acids as well as the fermentation process also effected the nutritional value. This meant that both air and sugar content affect the overall quality of a batch of silage and how long it takes to spoil (Kaiser et al. 2004).

Buffering capacity is perhaps one of the most important results from this experiment as it provides data on how stable a silage sample is (Moran, J 2005). It is clear from both Table 2 as well as all five graphs that buffering capacity was higher in the vacuumed samples. This is because these samples were able to produce more of the “good” acids (acetic and lactic) due to the fact that they could complete anaerobic fermentation without the addition of unwanted air or water (Moran, J 2005). This also proves that due to their higher buffering capacity, air-tight samples are less likely to spoil. The importance of buffering capacity can be shown in graphs 2 and 5 where there was a distinct difference in the speed of pH change between the non-vacuumed and vacuumed sample. The pH of the non-vacuumed samples changed much faster due to their lower buffering capacity and therefore lower stability (Moran, J 2005). The buffering capacity of each type of grass was highest when the added sugar content was above 0% and therefore this shows the importance of adding carbohydrates to silage as a stimulant for acid production.

The collected data shows that within the kikuyu samples, only one sample was close to being within the desired pH range, sample V3%. This is likely due to kikuyu being a tropical grass and therefore containing more moisture than a temperate grass and this is confirmed by the temperate grass samples being much closer to the desired pH range (Kaiser et al. 2004). This would suggest that the samples of kikuyu were not wilted thoroughly before being ensiled. Two ryegrass samples were measured at a pH of 4.3 which is quite close to the desired range (Ward & Ondarza 2008). These were the samples containing 2% and 3% sugar. In terms of pH, clover was the silage that stood out, with all of the vacuum packed clover silages within the desired range.

In terms of acid production, lactic acid is known as one of the major indicators of the success of a silage sample (Bernardes et al. 2018). This is shown in Table 2 where at the point where the pH of the sample is within or close to the desired range, lactic acid is at its highest percentage. Strong amounts of lactic acid production occurred in all sealed samples of silage with added sugar. This is because the additional levels of carbohydrates available for the anaerobic microorganisms to convert to lactic acid. In samples with low levels of lactic acid production, there was either not enough sugar added to for the production of high levels of lactic acid, or these samples were non-vacuumed and the presence of air did not provide the right conditions for anaerobic fermentation (Bernardes at al. 2018). Poor fermentation in a sample was shown by high levels of butyric acid (Zheng et al. 2018). Samples with higher than average butyric acid proportions included kikuyu samples; NV0%, NV3%, V0% and V1%, further highlighting the need for additional carbohydrates as well as an air-tight environment.

Nutritional value is also important when analysing silage samples. This is shown by the percentage of organic matter on a Dry Matter basis. Each sample’s organic matter content varied significantly depending on the levels of carbohydrate added (see Table 3). Within the kikuyu samples it was clear that the type of seal played the largest role in the amount of organic matter present. However, sugar content also played a role. The results show that at V0% organic matter content was at its highest (91.1%), this amount is lower at V1% and V3%, however, at V2% it climbs higher than V1% and V3%. Overall the organic matter results show lower amounts of added carbohydrate produce higher levels of organic matter. This means that more organic matter degradation occurred in the samples with the higher amounts of carbohydrate added in terms of thee kikuyu sample. This shows that the sample of V0% was most nutritious at 91.1%, just lower than the level of organic matter found in fresh grass (91.9%). It also shows that the addition of sugar to tropical grasses does not necessarily increase nutritional content. However, the fact that there is no pattern within the added sugar samples with V2% being an outlier provides some doubt on this theory. In fact, ryegrass and clover also each have an outlier in their organic matter results, however their data suggests that high carbohydrate content increases organic matter. This is backed up by research completed by Borreani et al. 2018 and may suggest the organic matter in kikuyu grass levels may be incorrect for this experiment.

In terms of ash content, it is clear from Table 3 that the lower organic matter samples had higher ash content. This means the nutritional value of the sample was reduced. Table 3 also shows the relationship between crude protein and ammonia nitrogen as a percentage of total nitrogen levels. High levels of ammonia were found in samples with a lower crude protein level. This is due to the breakdown of proteins as a result of poor fermentation and reduces the nutritional value of the sample as ruminants are not able to utilise the nutrient (Ward & Ondarza 2008).

From the analysis of results, it is clear that both the type of seal as well as the amount of added sugar play a role in the stability and nutrition of a silage sample (Moran, J 2005). The V3% kikuyu silage when compared to the other kikuyu silage samples (both vacuumed and non-vacuumed) was the stand-out. This is because it had the highest buffering capacity (and therefore the most stable pH), produced minimal amounts of butyric acid (as it was stored without contacting air), had the greatest levels of lactic acid due to the higher levels of added sugar. As well as this it still retained a relatively high level of organic matter along with crude protein. This means that not only is it the most stable silage, but it also retains nutrients well. However, in comparison to the ryegrass and clover samples, kikuyu still has improvements in the area of pH that must be made. An increase in added sugar to more than 3% would help the silage to produce more lactic acid, thus lowering the pH into the desired range and becoming more stable.

Conclusion

Overall, due to tropical grass having a much higher water content, wilting is an essential part of the ensiling process (Kaiser et al. 2004 & Moran, J 2005). The addition of sugars is also necessary to produce high quality silage in tropical grasses. This is because it helps promote lactic acid production and keep pH within a desired range.  Ensuring silage was vacuum sealed is also essential in the outcome of the silage as it produced an anaerobic environment which helped in lactic acid production rather than butyric acid production (Borreani et al. 2018). Due to the higher moisture content of the kikuyu samples, there was significant butyric acid production within these samples, however, the added sugar content was able to promote lactic acid production and therefore minimised the butyric acid content (Moran et al. 2005). With the addition of sugar, (perhaps more than the 3% used in the experiment), air-tight storage and a wilting process in place, it is likely tropical grasses such as kikuyu can be transformed into a high quality silage with significant nutritional benefits in livestock production. 

References

Bernardes, TF, Daniel, JLP, Adesogan, AT, McAllister, TA, Drouin, P, Nussio, LG, Huhtanen, P, Tremblay, GF, Belanger, G & Cai, Y 2018, ‘Silage review: Unique challenges of silages made in hot and cold regions’, Journal of Dairy Science, vol. 101, no. 5, pp. 4001-4009.

Borreani, G, Tabacco, E, Schmidt, RJ, Holmes, BJ & Muck, RE 2018, ‘Silage review: factors affecting dry matter and quality losses in silages’, Journal of Dairy Science, vol. 101, no. 5, pp. 3952-3979.

Gibson, T 1965, ‘Clostridia in Silage’, Journal of Applied Bacteriology, vol. 28, no. 1, viewed 8 May 2019, https://onlinelibrary-wiley-com.ezproxy.library.uq.edu.au/doi/abs/10.1111/j.1365-2672.1965.tb02126.x.

Hattersley, P 1983, ‘The distribution of C3 and C4 grasses in Australia in relation to climate’, Oecologia, vol. 57, no. 1-2, pp. 113-128.

Kaiser, AG, Piltz, JW, Burns, HM & Griffiths, NW 2004, Successful Silage, Dairy Australia and NSW Department of Primary Industries, Orange, NSW, viewed 9 May 2019, https://www.dairyaustralia.com.au/-/media/dairyaustralia/documents/farm/feedbase/supplements/successful-silage–04–silage-from-pastures-and-forage-crops.pdf?la=en&hash=FB15EC41CC84E22D94D52FE8C3055188E3C60234.

Moran, J 2005, Making Quality Silage, CSIRO and Department of Primary Industries, viewed 9 May 2019, http://www.publish.csiro.au/ebook/chapter/SA0501083.

Rodriguez, JA, Poppe, S & Meier, H 1989, ‘The influence of wilting on the quality of tropical grass silage in Cuba’, Archives of Animal Nutrition, vol. 39, no. 8-9, pp. 785-792.

Ward, RT & Ondarza, MB 2008, Fermentation analysis of silage: use and interpretation, Cumberland Valley Analytical services Inc., Hagerstown, MD, viewed 9May 2019, http://www.foragelab.com/media/fermentation-silage-nfmp-oct-2008.pdf.

Zheng, M, Dongze, N, Zuo, S, Mao, P, Meng, L & Xu, C 2017, ‘The effect of cultivar, wilting and storage period on fermentation and the clostridial community of alfalfa silage’, Italian Journal of Animal Science, vol. 17, no. 2, pp. 336-346.